Type 1 diabetes mellitus (T1D) is a chronic disease resulting from the autoimmune destruction of insulin- producing pancreatic beta cells. Although T1D is primarily associated with a dysfunctional immune response, increased access to human islet tissue has revealed surprisingly transient and heterogeneous levels of insulitis in T1D patients. Furthermore, recent genome-wide association studies (GWAS) have determined that a large number of T1D associated genes are expressed in pancreatic beta cells. Correspondingly, there is increasing indication of beta cell dysfunction occurring early in the T1D disease process and perhaps even in initiating the autoimmune response. There is also mounting evidence of diverse beta cell responses to the T-cell mediated immune attack, suggesting that beta cell stress and dysfunction may not only lead to beta cell death, but could possibly lead to increased beta cell plasticity and/or dedifferentiation. Understanding when and how these responses occur will not only identify potential new biomarkers of T1D, but will facilitate the development of nove therapies to prevent or block disease progression. To characterize the cellular and molecular events that are responsible for beta cell dysfunction and destruction early in the T1D process there is a need to develop novel genomics and proteomics technologies that can be used to evaluate available human tissues. Furthermore, given the heterogeneity associated with normal and genetically susceptible T1D islets, it will be critical to develop analytical tools that allow detailed characterization of human pancreatic tissues at single cell and near single cell resolution. In accordance with the goals of this RFA, the purpose of this grant is to use two powerful and complementary technology platforms - combFISH and nanoDESI - for in situ single cell resolution omics analyses of human pancreatic tissue that includes gene and protein expression, metabolite levels and lipid characterization to provide critical insights into T1D. The proposal brings together a strong team of investigators with relevant and complimentary areas of research expertise: Dr. Lori Sussel is a leader in the field of beta cell biology and Drs. Charls Ansong and Thomas Metz are experts in the field of proteomics and metabolomics. Dr. Julia Laskin is a leader in the field of imaging mass spectrometry and Dr. Galya Orr is an expert in quantitative super resolution and single-molecule fluorescence imaging. Dr. Mark Atkinson is a leader in T1D research. This combined expertise ensures the success of this research program. The goal of the proposal is to first optimize existing omics techniques on pancreatic islets derived from rodent models of T1D. The main thrust of the program will be to evaluate human normal and T1D islets and document and disseminate the data to the diabetes community. We will accomplish our goal through the following aims: (1) Technology Refinement in Rodent Models of T1D (2) Application of Technology to Human Islet Studies (3) Manage Data and Metadata to facilitate data integration and analysis.